Approximately 12% of the U.S. population or nearly 30,000,000 Americans have gallstones. This laboratory's long-term goal has been the elucidation of pathogenic mechanisms which may lead to new strategies to prevent gallstone formation. Key pathogenic factors in cholesterol gallstone formation include 1) cholesterol supersaturated bile, 2) gallbladder mucin hypersecretion, 3) cholesterol-phospholipid vesicle instability, 4) nucleation and growth of cholesterol monohydrate crystals, 5) diminished gallbladder motility, and 6) altered gallbladder mucosal function. In recent years, research efforts have focused on the events that lead to cholesterol crystal nucleation and/or growth. A number of pro- and anti- nucleating agents have been suggested. However, the key factors in this process have yet to be clearly defined. On the basis of preliminary human and animal data from this and other laboratories, we propose the following Central Hypothesis: Biliary nonmucin glycoproteins play a key role as pro- nucleating agents in cholesterol gallstone formation. Little is known, however, about the origin of nonmucin glycoproteins in either man or animal models. thus, the First Specific Aim will be to determine whether biliary nonmucin glycoproteins differ in serum hepatic bile, gallbladder bile, and the gallbladder's mucus gel. Recent data from this laboratory suggest that humans with cholesterol gallstones are more likely than controls to have an 84 kDa nonmucin glycoprotein in their gallbladder bile and four to six nonmucin glycoproteins in their stones which may be pro-nucleating agents. Moreover, several of the stone nonmucin glycoproteins have been found to be immunoglobulins. Therefore, the Second Specific Aim will be to determine which bile and stone nonmucin glycoproteins are pro-nucleating agents. We have been successful in producing monoclonal antibodies to pooled nonmucin glycoproteins from patients with pigment gallstones. More specific immunogens will be employed to produce monoclonal antibodies to pro- nucleating biliary nonmucin glycoproteins. Monoclonal antibody affinity chromatography will then be used to complete the Third Specific Aim which will be to purify and further characterize pro-nucleating biliary nonmucin glycoproteins. Clues to the early events of cholesterol crystal nucleation and growth may also be determined from the analysis of gallstones. Using electron microscopic and immunocytochemical techniques, the Fourth Specific Aim will be to identify which nonmucin glycoproteins are present at the core of cholesterol stones.